A cell culture system that enhances mononuclear cell IgE synthesis induced by recombinant human interleukin-4.
A new culture system is described in which recombinant human interleukin-4 (rhIL-4) consistently induces the synthesis of large quantities of IgE by human blood mononuclear cells (MNC). Unfractionated MNC were cultured in complete Iscove's modified Dulbecco's medium (C-IMDM), composed of IMDM enriched with human transferrin, bovine insulin, bovine serum albumin, oleic acid, palmitic acid, linoleic acid, and fetal calf serum (FCS). Under these culture conditions, MNC from four donors synthesized mean quantities of IgE of 76 ng/ml at plateau after stimulation with rhIL-4 in concentrations ranging from 0.04 to 80 ng/ml (plateau rhIL-4 concentrations were 5 ng/ml or greater). In contrast, rhIL-4 failed to induce significant IgE synthesis at any of those doses of rhIL-4 in parallel MNC cultures performed in RPMI 1640 supplemented with FCS (RPMI 1640). Additional optimal conditions for the induction of IgE synthesis in this system were a MNC concentration of 1-2 X 10(6)/ml and a culture time of 18 days. Variability was noted in the amount of IgE produced by different donors (CV 0.22) and by the same donor when tested on different occasions (mean CV 0.21), but no donor's MNC failed to produce significant IgE in response to rhIL-4 when cultured in C-IMDM. The geometric mean IgE production induced by optimal IL-4 concentrations for the entire group of 16 subjects was 36.8 ng/ml IgE, with the lowest day 18 mean IgE concentration for any donor being 10.6 ng/ml and the highest 372.2 ng/ml. The enhanced rhIL-4-induced IgE synthesis supported by C-IMDM was due to the combined effects of the added enrichment factors and not to differences in the viabilities of MNC cultured in C-IMDM and RPMI 1640. This culture system will alleviate the problems of inconsistent and low quantities of IgE induced by IL-4 that confound most current culture systems used to examine rhIL-4-induced IgE synthesis. It will, thereby, facilitate further investigation of the regulation of human IgE synthesis.
Claassen
J L
JL
Department of Pediatrics, Duke University School of Medicine, Durham, NC 27710.
Levine
A D
AD
Buckley
R H
RH
D000704:Analysis of Variance; D002478:Cells, Cultured / Q000187:drug effects / Q000378:metabolism*; D003470:Culture Media / Q000494:pharmacology; D006801:Humans; D007073:Immunoglobulin E / Q000096:biosynthesis*; D007074:Immunoglobulin G / Q000096:biosynthesis; D015847:Interleukin-4 / Q000494:pharmacology*; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000378:metabolism*; D008722:Methods; D011043:Pokeweed Mitogens / Q000494:pharmacology; D011994:Recombinant Proteins
D003470:Culture Media; D007074:Immunoglobulin G; D011043:Pokeweed Mitogens; D011994:Recombinant Proteins; D015847:Interleukin-4; D007073:Immunoglobulin E